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Journal of Biotechnology 202 (2015) 40–49

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Journal of Biotechnology

journal homepage: www.elsevier.com/locate/jbiotec

Discovery and development of Seliciclib. How systems biology approaches can lead to better drug performance

Hilal S. Khalila, Vanio Mitevb, Tatyana Vlaykovac, Laura Cavicchia, Nikolai Zheleva,∗

a CMCBR, SIMBIOS, School of Science, Engineering and Technology, Abertay University, Dundee DD1 1HG, Scotland, UK b Department of Chemistry and Biochemistry, Medical University of Sofia, 1431 Sofia, Bulgaria c Department of Chemistry and Biochemistry, Medical Faculty, Trakia University, Stara Zagora, Bulgaria

  • a r t i c l e i n f o
  • a b s t r a c t

Article history:

Received 10 August 2014 Received in revised form 26 February 2015 Accepted 27 February 2015 Available online 6 March 2015

Seliciclib (R-Roscovitine) was identified as an inhibitor of CDKs and has undergone drug development and clinical testing as an anticancer agent. In this review, the authors describe the discovery of Seliciclib and give a brief summary of the biology of the CDKs Seliciclib inhibits. An overview of the published in vitro and in vivo work supporting the development as an anti-cancer agent, from in vitro experiments to animal model studies ending with a summary of the clinical trial results and trials underway is presented. In addition some potential non-oncology applications are explored and the potential mode of action of Seliciclib in these areas is described. Finally the authors argue that optimisation of the therapeutic effects of kinase inhibitors such as Seliciclib could be enhanced using a systems biology approach involving mathematical modelling of the molecular pathways regulating cell growth and division.

Keywords:

Seliciclib Systems biology CDK

Crown Copyright © 2015 Published by Elsevier B.V. This is an open access article under the CC BY

license (http://creativecommons.org/licenses/by/4.0/).

1. Introduction

complex with its partner Cyclin B (CDK1/cyclin B), was required for prophase to metaphase transition, suggested that inhibitors of
The cell cycle is a fundamental biological process that is tightly regulated by the activity of a series of kinases termed the CyclinDependent Kinases (CDKs). These are such named because of the requirement for binding CDK specific cyclins for their activity (Gran˜a and Reddy, 1995). The activities of these kinases must follow a specific sequence to allow normal cell cycle progression (Morgan, 1997) and abberations in the control of the cell cycle have been linked to a variety of diseases including cancer, inflammatory conditions and neurodegenerative disorders (Zhivotovsky and Orrenius, 2010). The cell cycle proceeds through various checkpoints each of which is regulated by the activity of CDKs that are in turn, regulated by signalling pathways either promoting or inhibit-

ing cell (Chiarle et al., 2001).

The first CDK to be discovered was CDK1, which was originally identified in starfish oocytes as “Maturation Promoting Factor” or MPF. It was found that when oocytes previously arrested in the prophase of the cell cycle, were injected with CDK1, this caused their entry into metaphase, a process known to be associated with protein phosphorylation (Meijer and Guerrier, 1984; Labbé et al., 1989). This observation, that the activity of CDK1, in this kinase could be useful in the treatment of proliferative disor-

ders (Pondaven et al., 1990; Rialet and Meijer, 1991). Supporting

this hypothesis, Dimethylaminopurine (DMAP), a drug that was initially identified as a potent inhibitor of mitosis in sea urchins (Rebhun et al., 1973) was subsequently shown to exert its action through inhibition of CDK1/cyclin B complex (Rialet and Meijer,

1991; Neant and Guerrier, 1988). DMAP and a related purine

isopentyladenine had in vitro IC50 values of 120 ␮M and 55 ␮M against CDK1/cyclin B respectively. The fact that isopentyadenine was an intermediate in the biosynthesis of the cytokinin group of plant hormones led to a collaboration between Laurent Meijer of the Biological Station in Roscoff and Jaroslav Vesely and Miroslav Strnad at the Institute of Experimental Botany in Olomouc in the Czech Republic. Their collaborative work resulted in the synthesis of a number of substituted purine molecules, the most promising of which was 2-(2-hydroxyethylamino)-6- benzylamino-9-methylpurine. This molecule, which was named Olomoucine, was specific in its inhibitory action towards CDK and MAPK (Vesely´ et al., 1994), an observation which, at the time, was surprising for an ATP analogue. Olomoucine was considerably more potent with an IC50 value of 7 ␮M against CDK1/cyclin B in vitro. Strnad in collaboration with Michel Legraverend of the Institute Marie Curie at Orsay worked together to synthesise more potent and more specific substituted purines, the best of

∗

Corresponding author. Tel.: +44 1382308536.

E-mail address: [email protected] (N. Zhelev). http://dx.doi.org/10.1016/j.jbiotec.2015.02.032

0168-1656/Crown Copyright © 2015 Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

H.S. Khalil et al. / Journal of Biotechnology 202 (2015) 40–49

41

Table 1

these kinases is required for initiation and progression of cellular division chemical inhibition of the CDKs has the potential to be useful in proliferative diseases such as cancer.

Studies demonstrating CDK inhibition by Roscovitine in vitro and in vivo.

  • CDK/Cyclin type
  • Studied model
  • Reference

CDK1

CDK1

Lung cancer cell line

Human Colorectal cancer cell line

Schutte et al. (1997) Abal et al. (2004)

2. CDK1

CDK1/Cyclin B CDK1/Cyclin B
Xenopus oocytes In vitro kinase assay

Meijer et al. (1997) Meijer et al. (1997), Raynaud et al. (2005) Meijer et al. (1997), Raynaud et al. (2005) Iseki et al. (1997)

CDK1, also referred to as the mitotic kinase, forms a complex with cyclin B (Malumbres and Barbacid, 2007) At 297 amino acids in length and with a molecular weight of 34 kDa its activity is modulated by post-translational modification, being activated or inhibited by site-specific phosphorylation by regulatory kinases including Wee1, Mik1 amd Myt1 on Threonine 161,Tyrosine 15 or Threonine 14 (Schafer, 1998). Hyperactivity of CDK1 either through overexpression of Cyclin B1 or hyperphosphoryation of CDK1 has been observed, observed in several tumours, including breast-, colon- and prostate carcinoma (Pérez de Castro et al., 2007) this supporting the hypothesis that dysregulation of this kinases could cause uncontrolled cellular division.

  • CDK1/Cyclin B
  • In vitro kinase assay

CDK1, CDK2 CDK2 CDK2
Human Gastric cell lines Human Pancreatic cell line Human Osteosarcoma, Cervical, Lung carcinoma cell lines

Iseki et al. (1998) Zhang et al. (2004a,b)

CDK2/Cyclin A, E
& B
In vitro kinase assay

Meijer et al. (1997), Havlícek et al. (1997), Biglione et al. (2007), Raynaud et al. (2005) Meijer et al. (1997)

CDK2/Cyclin B CDK2, Cyclin E
Mouse lymphocytic leukaemia cell line In vitro kinase assay, human tumour cell lines, mouse model

McClue et al. (2002)

3. CDK2

  • CDK2/Cyclin B
  • HCT116 colon cancer cell

line Human breast cancer cell lines

Raynaud et al. (2005) Nair et al. (2011)

Dysregulation of CDK2 activity has also been observed in a variety of malignancies further supporting the theory that inhibition of the CDKs be Roscovitine could be beneficial in the treatment of proliferative diseases. Although CDK2 is a key cell cycle regulator, critical for the transition into the S-phase of the cell cycle, mice lacking the kinase are viable, suggesting that there are other kinases which can compensate for any lack in CDK2 activity (Berthet et al., 2003). CDK2 activity is controlled not just by phosphorylation events by complexation with inhibitory protein partners such as Cip/Kip and of course its cyclin partners Cyclin E and Cyclin A dysregulation of which has been observed in malignancies (Pérez de

Castro et al., 2007).

CDK2/Cyclin D1, Cyclin A2

  • CDK4/Cyclin D1
  • In vitro kinase assay

Meijer et al. (1997), Raynaud et al. (2005) Raynaud et al. (2005)

  • CDK4/Cyclin D1
  • HCT116 colon cancer cell

line In vitro kinase assay In vitro kinase assay
CDK5/P35 CDK6/Cyclin D3

Meijer et al. (1997) Meijer et al. (1997), Raynaud et al. (2005) Raynaud et al. (2005) Biglione et al. (2007), Raynaud et al. (2005)

CDK7/Cyclin H CDK9/Cyclin T1
Invitro kinase assay In vitro kinase assay & Hela cells

which, 6-(benzylamino)-2(R)-[[1-(hydroxymethyl)propyl]amino]- 9-isopropylpurine, termed Roscovitine, had an in vitro IC50 value of 0.45 ␮M against the CDK1/cylin B complex (Havlícek et al., 1997). Roscovitine and olomoucine were subsequently co-crystallised with CDK2 and these structures were used as the basis of molecular models for guiding further medicinal chemistry programmes (De

Azevedo et al., 1997).

Roscovitine has been demonstrated to be a potent inhibitor of a number of CDKs including CDK1/cyclin B (0.65 ␮M), CDK2/cyclin A (0.7 ␮M), CDK2/cyclin E (0.7 ␮M), CDK5/p35 (0.2 ␮M), CDK7/cyclin H (0.49 ␮M), and CDK9/cyclin H (0.79 ␮M). However, because Roscovitine is an ATP competitive molecule, the precise IC50 values reported vary depending on the concentration of ATP used in the

in vitro assay (Wang and Fischer, 2008; Meijer et al., 1997; McClue et al., 2002; Biglione et al., 2007). CDK4/cyclin D1, CDK6/cyclin D3

and over 80 other kinases tested were all insensitive or only weakly inhibited by Roscovitine (Bain et al., 2003, 2007).
As an inhibitor of CDKs 1, 2, 5, 7 and 9 Roscovitine can impact a variety of cellular functions in tissue dependent manner. A summary of studies, demonstrating CDK inhibition by Roscovitine is shown in Table 1. Thus, it is important to have knowledge of individual CDK functions especially while employing a broad spectrum CDK inhibitor. Here, we will briefly examine the biology of different CDKs in an effort to ascertain which therapeutic areas inhibitors of these kinases could impact upon.

4. CDK5

CDK5 is required for central and peripheral nervous system function (Cruz and Tsai, 2004) and has been implicated in numerous neuronal functions including cytoarchitecture in the brain, neuronal migration, synaptic plasticity, learning and memory and may be involved in the development of neurodegenerative disorders including Alzheimer’s and Parkinson’s Diseases (Angelo et al.,

2006; Cruz and Tsai, 2004; Dhavan and Tsai, 2001). Treatment of

the lower eukaryote Dictyostelium discoideum with Roscovitine led to an inhibition not only of the single-cell growth phase of the organism but also arrested translocation of the protein between the nucleus and cytoplasm raising the possibility that at least part of the biological effects of Roscovitine may be due to secondary effects such as protein location (Huber and O’Day, 2012). Inhibition of CDK5 in lower eukaryotes has also indicated a role for the kinase in development, cytoskeletal organisation and calcium

channel function (Huber and O’Day, 2012; Prithviraj et al., 2012;

Wen et al., 2013). CDK5 has also been implicated in modulating the metastatic potential of breast and prostate carcinomas (Goodyear

and Sharma, 2007; Strock et al., 2006). It is unusual in that it

exhibits kinase activity only when bound to non-cyclin activators CDK5R1 and CDK5R2 although structural studies on these proteins have shown structural similarity with the cyclins (Cheung and Ip,

2012).

As an inhibitor of the CDK family roscovitine can potentially impact upon a number of fundamental processes in cellular biology. Cell division has to be highly regulated and is an area of cellular biology in which the CDK family is heavily involved. Roscovitinetarget CDKs 1 and 2 are involved in the control of the transition of cells from G2 to M and G1 to S respectively and as the activity of
These observations that CDK5, a kinase that is inhibited by
Roscovitine broaden the therapeutic applications of the compound further beyond the less well documented areas of proliferative disease and virology. The potential of Roscovitine to treat neurological disorders such as Alzheimers and Parkinson’s is very exciting given the paucity of treatments currently available for these treatments

42

H.S. Khalil et al. / Journal of Biotechnology 202 (2015) 40–49

and its low toxicity and excellent tolerability are surely plus points in this therapeutic area. kinetics of the cell cycle in the human lung cancer cell line MR65 and the neuroblastoma line CHP212 (Schutte et al., 1997). In this study, cells exposed to either of the compounds showed delays in the transitions from G1 to S phase and from G2/M to G1 as well as a prolonged S phase. They also observed changes in cell morphology that were indicative of apoptosis in the treated cells. In a study using normal human fibroblasts, Alessi et al. reported a reversible block in G1 after Roscovitine or Olomoucine treatments (Alessi et al., 1998) and reduced levels of hyper-phosphorylated Rb, indicating cell cycle arrest, but unchanged levels of Proliferating Cell Nuclear Antigen (PCNA) and Cyclins D1 and E. In another study, treatment of human gastric cell lines SIIA, AGS, MKN45- 630 and SNU-1, resulted in an increase in the proportion of cells in G2/M and S phases. In SIIA cells, treatment led to a reduction in levels of phosphorylated Histone H1, suggesting that the compound was inhibiting CDK1 and CDK2 (Iseki et al., 1997). A year later, same group also examined four human pancreatic cell lines with differing genetic lesions and showed that Roscovitine and Olomoucine inhibited CDK2 activity and cellular proliferation independent of the p53, K-Ras or p16 status (Iseki et al., 1998). During the same year, Mgbonyebi and colleagues investigated the effect of Roscovitine on the proliferation of immortal and neoplastic breast cancer cells and reported that Oestrogen Receptor (ER) positive and ER negative cell lines were sensitive to Roscovitine (Mgbonyebi et al., 1998). In a further study, the same group reported that treatment of ER-ve MD-MB-231 cells with Roscovitine for between

5. CDK7

CDK7 binds not only to its cyclin partner Cyclin H but also forms a trimer with a third partner, MAT1. Termed the Cyclin-dependent kinase Activating Kinase or CAK this trimer phosphorylates CDKs 1, 2, 4 and 6 on their key activating residues (Lolli and Johnson, 2005). A role in cell division has been observed in some eukaryotic systems including yeast, where loss of activity causes cell cycle arrest and drosophila in which mutations are lethal before or during pupation

(Larochelle et al., 1998; Wallenfang and Seydoux, 2002). In mam-

malian cells loss of MAT1 induces cellular arrest in G1 and cell death by apoptosis (Wu et al., 1999). The cell-cycle role of CDK7 in cell death is less clear cut than some of the other CDKs due to the fact it is also involved in the control of transcriptional. CAK forms part of the large multimeric general transcription factor TFIIH where it phosphorylates the C-Terminal Domain (CTD) of RNA Polymerase II improving the efficiency of transcriptional initiation and elongation (Maldonado and Reinberg, 1995). As the kinase has multiple biological effects it is more difficult to define unambiguously which inhibition causes which effect.

6. CDK9

  • 1
  • and 10 days induced morphological changes in the cells

CDK9 with its partner Cyclins T or K also forms part of the

transcriptional machinery being a core part of the multi-subunit positive transcription elongation factor b (p-TEFb) (Loyer et al.,

2005; Malumbres and Barbacid, 2005; Romano and Giordano,

2008; Yu and Cortez, 2011) which is involved in improving the transcriptional elongation from RNA Pol II dependent promoters. This class of promoter drives expression of multiple key developmental and cellular response genes as well as the majority of protein

encoding genes (Nechaev and Adelman, 2011).

The discovery of role of the CDKs 7 and 9 in the control of gene transcription opened up new possibilities for roscovitine in new therapeutic areas, most importantly in virology where the importance of their activity has been recognised in the replication of Herpes Simplex Virus, Human Imunodeficiency Virus and Human

Cytomegalovirus (Boeing et al., 2010; Durand and Roizman, 2008; Schang et al., 1998; Yang et al., 1997)

consistent with the induction of apoptosis (Mgbonyebi et al.,

1999).

Responses to Roscovitine have also been investigated in combination with a number of other chemotherapeutic agents in vitro. It has been shown to have potential synergistic relationships with camptothecin in the breast tumour line MCF7, the histone deacetylase inhibitor LAQ824 in leukaemic cell lines HL60, with doxorubicin in sarcoma cell lines and also with irinotecan in a p53-

mutated colon cancer (Lu et al., 2001; Lambert et al., 2008; Abal et al., 2004).

We have previously studied the effects of R-Roscovitine
(CYC202) on the physiology of normal and transformed human cells. These studies revealed for the first time that at therapeutic doses, the drug is not toxic to normal keratinocytes, but at higher doses CYC202 can affect components of major signalling pathways, (e.g. p38), highlighting potential side-effects of the drug in vivo (Atanasova et al., 2005, 2007). In addition to the induction of apoptosis and cell cycle effects, Roscovitine has been reported to inhibit DNA synthesis in primary human glioma samples by almost 90% (Yakisich et al., 1999) as well as inducing mucinous differentiation in the human non-small cell lung cancer line NCI-H348 (Lee et al.,

1999).

In summary Roscovitine has been reported to induce apoptosis in several cell lines independently of p53 status. Cell death has been detected in all phases of the cell cycle via a variety of potential mechanisms including inhibition of the cell cycle and effects on transcription due to reduced phosphorylation of the CTD of RNA polymerase II by CDK7 and CDK9 (WesierskaGadek et al., 2005, 2008). However, treatment with Roscovitine had relatively little impact on global transcription with only a small number of transcripts found to be significantly reduced. It is worth noting that those proteins whose transcript level was found to be reduced by Roscovitine treatment were mostly prosurvival factors on which tumour cells may be more dependent than normal cells (Meijer and Galons, 2006). These observations suggest that cell death induced by Roscovitine may be due to the reduction in levels of a small number of survival factors such as

Mcl-1, XIAP and survivin (Lacrima et al., 2005; Mohapatra et al., 2005).

7. In vitro studies of Roscovitine as an anti cancer drug

Although CDKs play pivotal roles in a range of cellular functions, studies with Roscovitine have focussed largely on its inhibitory effects on cell cycle progression, mainly with a view to its development as a potential anti-cancer agent. Roscovitine has been tested on more than 100 cell lines, including the NCI-60 panel of the United States’ National Cancer Institute (Shoemaker, 2006).
In 1997, Meijer et al. showed that constant exposure to Roscovitine over a 48 h period inhibited the growth of 60 different cell lines from 9 different tissue types when compared with non-exposed cells. The average IC50 across all cell lines was 16 ␮M (Meijer et al., 1997). In a separate study Raynaud and colleagues reported that Roscovitine inhibited the growth of 24 cell lines with an average IC50 value of 14.6 ␮M (Raynaud et al., 2005). Other studies have shown that in the mouse leukaemia cell line L1210, Roscovitine led to an accumulation of the cells in G2/M cycle. This accumulation of cells in the G2/M phase was also observed in A549 human lung cancer cell lines in a detailed study by McClue et al. (2002). This group demonstrated that 24 h of treatment with Roscovitine led to a significant increase in apoptosis. Schutte and colleagues examined the effects of both roscovitine and olomoucine on the

H.S. Khalil et al. / Journal of Biotechnology 202 (2015) 40–49

43

8. In vivo studies of Roscovitine as an anti cancer drug

56% clearly showing that these two compounds act synergistically to reduce tumour growth (Fleming et al., 2008).
Roscovitine has been tested extensively in animal models, largely in xenograft models of various cancers, as part of its development as an anticancer agent. In addition it has been examined in all the standard toxicology tests required by regulatory authorities and although the authors are not aware of any toxicological issues, that data will not be discussed in this review.
In another study, Roscovitine was tested in a xenograft GBM43 glioma model in combination with an experimental PI-3 kinase inhibitor, PIK-90 (Cheng et al., 2012). Both drugs were dosed intraperitoneally four times per day for 12 days, Roscovitine at 50 mg/kg and PIK-90 at 40 mg/kg. At the end of the 12-day treatment period, mice treated with the combination of the drugs showed 75% reduction in tumour volume as compared to that in untreated animals. Treatment with Roscovitine or PIK-90 alone reduced tumour volumes by approximately 40% and 50% respectively indicating that the combination of the two drugs was not more than additive

(Cheng et al., 2012).

We have identified the specific CDK inhibitor, p27 and its substrate Rb, as biomarkers for CYC202 mediated cell growth inhibition, and demonstrated its usefulness for monitoring inhibition of its major target (CDK2) in cellulo and in vivo (Whittaker et al.,

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  • Roscovitine in Cancer and Other Diseases

    Roscovitine in Cancer and Other Diseases

    Review Article Page 1 of 12 Roscovitine in cancer and other diseases Jonas Cicenas1,2,3, Karthik Kalyan2,4, Aleksandras Sorokinas2, Edvinas Stankunas2,5, Josh Levy2,6, Ingrida Meskinyte7, Vaidotas Stankevicius2,8,9, Algirdas Kaupinis3, Mindaugas Valius3 1CALIPHO Group, Swiss Institute of Bioinformatics, Geneva, Switzerland; 2MAP Kinase Resource, Bern, Switzerland; 3Proteomics Centre, Vilnius University Institute of Biochemistry, Vilnius, Lithuania; 4Systems Biomedicine Division and Department of Virology and Immunology, Haffkine Institute for Training Research and Testing, Mumbai, India; 5Department of Biochemistry, Vilnius University, Vilnius, Lithuania; 6RTI International, Research Triangle Park, NC, USA; 7Lithuanian Centre of Non-Formal Youth Education Vilnius, Lithuania; 8National Cancer Institute, Vilnius, Lithuania; 9Vilnius University, Vilnius, Lithuania Correspondence to: Jonas Cicenas. Swiss Institute of Bioinformatics, CALIPHO Group, CMU-1, rue Michel Servet’ CH-1211, Geneva 4, Switzerland. Email: [email protected]. Abstract: Roscovitine [CY-202, (R)-Roscovitine, Seliciclib] is a small molecule that inhibits cyclin-dependent kinases (CDKs) through direct competition at the ATP-binding site. It is a broad-range purine inhibitor, which inhibits CDK1, CDK2, CDK5 and CDK7, but is a poor inhibitor for CDK4 and CDK6. Roscovitine is widely used as a biological tool in cell cycle, cancer, apoptosis and neurobiology studies. Moreover, it is currently evaluated as a potential drug to treat cancers, neurodegenerative diseases, inflammation, viral infections, polycystic kidney disease and glomerulonephritis. This review focuses on the use of roscovitine in the disease model as well as clinical model research. Keywords: Cyclin-dependent kinases (CDK); small molecule inhibitor; roscovitine; cancer; neurodegeneration; kidney diseases Submitted Dec 16, 2014. Accepted for publication Mar 16, 2015.
  • In Combination with Cytotoxic Agents in Human Uterine Sarcoma Cell Lines

    In Combination with Cytotoxic Agents in Human Uterine Sarcoma Cell Lines

    ANTICANCER RESEARCH 27: 273-278 (2007) Seliciclib (CYC202; r-Roscovitine) in Combination with Cytotoxic Agents in Human Uterine Sarcoma Cell Lines HELEN M. COLEY, CHRISTINE F. SHOTTON and HILARY THOMAS Postgraduate Medical School, University of Surrey, Guildford, Surrey GU2 7WG, U.K. Abstract. Background: Inhibition of cyclin-dependent kinases the cell cycle as an approach to treat cancer. Seliciclib (CDKs) has recently emerged as an interesting approach to treat (CYC202), the r-enantiomer of the cell cycle inhibitory human malignancies. This was explored in human leiomyo- agent roscovitine has been developed as a potent CDK2 sarcoma (LMS) lines, which represent a tumour associated with inhibitor and is currently in phase II clinical trials. poor survival, chemo-unresponsiveness and deregulation of cell Preclinical studies involving other CDK inhibitors, such as cycle components. Materials and Methods: Using isobologram flavopiridol have demonstrated their interaction with a analysis with MTT chemosensitivity testing, the effects of the CDK number of different cytotoxic agents in a synergistic manner inhibitor seliciclib (CYC202, R-roscovitine) when used alone or (5). We have explored this approach by examining the in combination with paclitaxel was studied in uterine cancer cell effects of seliciclib combined with paclitaxel in three human lines. Apoptotic endpoints were also examined via Annexin V uterine sarcoma cell line models in terms of any synergy and assay using flow cytometry and Western blotting. Results: Overall effects on apoptosis. seliciclib combined with paclitaxel proved synergistic for all cell lines. This was concomitant with an enhanced apoptotic effect Materials and Methods and downregulation of the IAP survivin. Conclusion: Our data support the use of seliciclib as part of combination therapy for Chemicals and reagents.
  • Phenotype-Based Drug Screening Reveals Association Between Venetoclax Response and Differentiation Stage in Acute Myeloid Leukemia

    Phenotype-Based Drug Screening Reveals Association Between Venetoclax Response and Differentiation Stage in Acute Myeloid Leukemia

    Acute Myeloid Leukemia SUPPLEMENTARY APPENDIX Phenotype-based drug screening reveals association between venetoclax response and differentiation stage in acute myeloid leukemia Heikki Kuusanmäki, 1,2 Aino-Maija Leppä, 1 Petri Pölönen, 3 Mika Kontro, 2 Olli Dufva, 2 Debashish Deb, 1 Bhagwan Yadav, 2 Oscar Brück, 2 Ashwini Kumar, 1 Hele Everaus, 4 Bjørn T. Gjertsen, 5 Merja Heinäniemi, 3 Kimmo Porkka, 2 Satu Mustjoki 2,6 and Caroline A. Heckman 1 1Institute for Molecular Medicine Finland, Helsinki Institute of Life Science, University of Helsinki, Helsinki; 2Hematology Research Unit, Helsinki University Hospital Comprehensive Cancer Center, Helsinki; 3Institute of Biomedicine, School of Medicine, University of Eastern Finland, Kuopio, Finland; 4Department of Hematology and Oncology, University of Tartu, Tartu, Estonia; 5Centre for Cancer Biomarkers, De - partment of Clinical Science, University of Bergen, Bergen, Norway and 6Translational Immunology Research Program and Department of Clinical Chemistry and Hematology, University of Helsinki, Helsinki, Finland ©2020 Ferrata Storti Foundation. This is an open-access paper. doi:10.3324/haematol. 2018.214882 Received: December 17, 2018. Accepted: July 8, 2019. Pre-published: July 11, 2019. Correspondence: CAROLINE A. HECKMAN - [email protected] HEIKKI KUUSANMÄKI - [email protected] Supplemental Material Phenotype-based drug screening reveals an association between venetoclax response and differentiation stage in acute myeloid leukemia Authors: Heikki Kuusanmäki1, 2, Aino-Maija
  • ABCB1 As Predominant Resistance Mechanism in Cells with Acquired SNS-032 Resistance

    ABCB1 As Predominant Resistance Mechanism in Cells with Acquired SNS-032 Resistance

    www.impactjournals.com/oncotarget/ Oncotarget, Vol. 7, No. 36 Research Paper ABCB1 as predominant resistance mechanism in cells with acquired SNS-032 resistance Nadine Löschmann1,*, Martin Michaelis2,*, Florian Rothweiler1, Yvonne Voges1, Barbora Balónová3, Barry A. Blight3, Jindrich Cinatl Jr1 1Institut für Medizinische Virologie, Klinikum der Goethe-Universität, 60596 Frankfurt am Main, Germany 2Centre for Molecular Processing and School of Biosciences, University of Kent, Canterbury, UK 3School of Physical Sciences, University of Kent, Canterbury, UK *These authors equally contributed to this work Correspondence to: Jindrich Cinatl Jr, email: [email protected] Keywords: ABCB1, CDK inhibitor, multi-drug resistance, neuroblastoma, cancer Received: February 19, 2016 Accepted: July 27, 2016 Published: August 09, 2016 ABSTRACT The CDK inhibitor SNS-032 had previously exerted promising anti-neuroblastoma activity via CDK7 and 9 inhibition. ABCB1 expression was identified as major determinant of SNS-032 resistance. Here, we investigated the role of ABCB1 in acquired SNS-032 resistance. In contrast to ABCB1-expressing UKF-NB-3 sub- lines resistant to other ABCB1 substrates, SNS-032-adapted UKF-NB-3 (UKF-NB- 3rSNS- 032300nM) cells remained sensitive to the non-ABCB1 substrate cisplatin and were completely re-sensitized to cytotoxic ABCB1 substrates by ABCB1 inhibition. Moreover, UKF-NB-3rSNS-032300nM cells remained similarly sensitive to CDK7 and 9 inhibition as UKF-NB-3 cells. In contrast, SHEPrSNS-0322000nM, the SNS-032-resistant sub-line of the neuroblastoma cell line SHEP, displayed low level SNS-032 resistance also when ABCB1 was inhibited. This discrepancy may be explained by the higher SNS-032 concentrations that were used to establish SHEPrSNS-0322000nM cells, since SHEP cells intrinsically express ABCB1 and are less sensitive to SNS-032 (IC50 912 nM) than UKF-NB-3 cells (IC50 153 nM).
  • Full Text (PDF)

    Full Text (PDF)

    Published OnlineFirst April 2, 2015; DOI: 10.1158/1078-0432.CCR-14-0959 Molecular Pathways Clinical Cancer Research Molecular Pathways: Leveraging the BCL-2 Interactome to Kill Cancer Cells—Mitochondrial Outer Membrane Permeabilization and Beyond Hetal Brahmbhatt1,2, Sina Oppermann2, Elizabeth J. Osterlund2,3, Brian Leber4, and David W. Andrews1,2,3 Abstract The inhibition of apoptosis enables the survival and prolif- which antagonizes the activity of BCL-2, is currently the furthest eration of tumors and contributes to resistance to conventional in clinical trials and shows promising activity in many lym- chemotherapy agents and is therefore a very promising avenue phoid malignancies as a single agent and in combination with for the development of new agents that will enhance current conventional chemotherapy agents. Here, we discuss strategies cancer therapies. The BCL-2 family proteins orchestrate apo- to improve the specificity of pharmacologically modulating ptosis at the mitochondria and endoplasmic reticulum and are various antiapoptotic BCL-2 family proteins, review additional involved in other processes such as autophagy and unfolded BCL-2 family protein interactions that can be exploited for the protein response (UPR) that lead to different types of cell death. improvement of conventional anticancer therapies, and high- Over the past decade, significant efforts have been made light important points of consideration for assessing the activ- to restore apoptosis using small molecules that modulate the ity of small-molecule BCL-2 family protein modulators. Clin activity of BCL-2 family proteins. The small molecule ABT-199, Cancer Res; 21(12); 2671–6. Ó2015 AACR. Background are predominantly involved in the intrinsic pathway, in which they regulate mitochondrial outer membrane permeabilization Tumorigenesis is a complex multistep process that occurs when (MOMP).
  • View a Copy of This Licence, Visit

    View a Copy of This Licence, Visit

    Cole et al. Trials (2021) 22:433 https://doi.org/10.1186/s13063-021-05384-5 STUDY PROTOCOL Open Access TRAFIC: statistical design and analysis plan for a pragmatic early phase 1/2 Bayesian adaptive dose escalation trial in rheumatoid arthritis M. Cole1, C. Yap2, C. Buckley3,W.F.Ng4, I. McInnes5, A. Filer3, S. Siebert5, A. Pratt4, J. D. Isaacs4 and D. D. Stocken6* Abstract Background: Adaptive model-based dose-finding designs have demonstrated advantages over traditional rule- based designs but have increased statistical complexity but uptake has been slow especially outside of cancer trials. TRAFIC is a multi-centre, early phase trial in rheumatoid arthritis incorporating a model-based design. Methods: A Bayesian adaptive dose-finding phase I trial rolling into a single-arm, single-stage phase II trial. Model parameters for phase I were chosen via Monte Carlo simulation evaluating objective performance measures under clinically relevant scenarios and incorporated stopping rules for early termination. Potential designs were further calibrated utilising dose transition pathways. Discussion: TRAFIC is an MRC-funded trial of a re-purposed treatment demonstrating that it is possible to design, fund and implement a model-based phase I trial in a non-cancer population within conventional research funding tracks and regulatory constraints. The phase I design allows borrowing of information from previous trials, all accumulated data to be utilised in decision-making, verification of operating characteristics through simulation, improved understanding for management and oversight teams through dose transition pathways. The rolling phase II design brings efficiencies in trial conduct including site and monitoring activities and cost.
  • Synergistic Inhibition of Erbb Signaling by Combined Treatment

    Synergistic Inhibition of Erbb Signaling by Combined Treatment

    Cancer Therapy: Preclinical Synergistic Inhibition of ErbB Signaling by Combined Treatment with Seliciclib and ErbB-Targeting Agents IanN.Fleming,MoragHogben,SheelaghFrame,StevenJ.McClue,andSimonR.Green Abstract Purpose: The aims of this studywere to investigate whether the cyclin-dependent kinase inhi- bitor seliciclib could synergize with agents that target ErbB receptors and to elucidate the mole- cular mechanism of the observed synergy. Experimental Design: Synergy between seliciclib and ErbB receptor targeted agents was in- vestigated in various cell lines using the Calcusyn median effect model.The molecular mechanism of the observed synergy was studied in cultured cells, and the combination of seliciclib and the epidermal growth factor receptor (EGFR) inhibitor erlotinib was evaluated in an H358 xenograft model. Results: Seliciclib synergized with the anti-HER2 antibody trastuzumab in a breast cancer cell line, which overexpresses the HER2 receptor, and with the erlotinib analogue AG1478 in non ^ small cell lung cancer cell lines. In the H358 non ^ small cell lung cancer cell line, synergy involved decreased signaling from the EGFR, with AG1478 directlyinhibiting kinase activitywhile seliciclib decreased the levels of keycomponents of the receptor signaling pathway,resulting in enhanced loss of phosphorylated extracellular signal-regulated kinase and cyclin D1.The combination of seliciclib and erlotinib was evaluated further in an H358 xenograft and shown to be significantly more active than either agent alone. An enhanced loss of cyclin D1was also seen in vivo. Conclusions: This is the first report that investigates combining seliciclib with an EGFR inhibitor. The combination decreased signaling from the EGFR in vitro and in vivo and was effective in cell lines containing either wild-type or mutant EGFR, suggesting that it may expand the range of tumors that respond to erlotinib, and therefore, such combinations are worth exploring in the clinic.
  • Review Article Cell Cycle Inhibition Without Disruption of Neurogenesis Is a Strategy for Treatment of Aberrant Cell Cycle Diseases: an Update

    Review Article Cell Cycle Inhibition Without Disruption of Neurogenesis Is a Strategy for Treatment of Aberrant Cell Cycle Diseases: an Update

    The Scientific World Journal Volume 2012, Article ID 491737, 13 pages The cientificWorldJOURNAL doi:10.1100/2012/491737 Review Article Cell Cycle Inhibition without Disruption of Neurogenesis Is a Strategy for Treatment of Aberrant Cell Cycle Diseases: An Update Da-Zhi Liu and Bradley P. Ander Department of Neurology and the MIND Institute, University of California at Davis, Sacramento, CA 95817, USA Correspondence should be addressed to Da-Zhi Liu, [email protected] Received 13 October 2011; Accepted 17 November 2011 Academic Editors: F. Bareyre and B. K. Jin Copyright © 2012 D.-Z. Liu and B. P. Ander. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Since publishing our earlier report describing a strategy for the treatment of central nervous system (CNS) diseases by inhibiting the cell cycle and without disrupting neurogenesis (Liu et al. 2010), we now update and extend this strategy to applications in the treatment of cancers as well. Here, we put forth the concept of “aberrant cell cycle diseases” to include both cancer and CNS diseases, the two unrelated disease types on the surface, by focusing on a common mechanism in each aberrant cell cycle reentry. In this paper, we also summarize the pharmacological approaches that interfere with classical cell cycle molecules and mitogenic pathways to block the cell cycle of tumor cells (in treatment of cancer) as well as to block the cell cycle of neurons (in treatment of CNS diseases). Since cell cycle inhibition can also block proliferation of neural progenitor cells (NPCs) and thus impair brain neurogenesis leading to cognitive deficits, we propose that future strategies aimed at cell cycle inhibition in treatment of aberrant cell cycle diseases (i.e., cancers or CNS diseases) should be designed with consideration of the important side effects on normal neurogenesis and cognition.